Stroke and the resulting neurological deficits are the most common cause for permanent disability in the United States. The death of neurons by apoptosis and the loss of axons and dendrites play a major role in damage in the penumbra of a stroke and in many neurodegenerative diseases. The aim of the proposed research project is to elucidate the mechanism of a synergistic effect of combinatorial treatment with erythropoietin (EPO) and insulin-like growth factor I (IGF-I) in order to ameliorate the acute and long-term disabilities after stroke. Detailed behavioral studies, including tests of cognitive, locomotor and sensorimotor function, will be conducted in groups of mice. Neurological scores of mice in subgroups subjected to experimental stroke and treated with each cytokine alone will be compared to those administered EPO+IGF-I or vehicle. Neurological benefits of each treatment will be based on the combination of these scores. The occurrence of stroke, indicated by loss of neurons and their extensions in histological sections, and the possible beneficiary effect of EPO?I will be documented by measuring stroke volume. In addition, we will study the signaling pathway triggered by EPO and IGF-I interaction with their respective receptors (EPO-R and IGF-IR) in neurons. We will investigate the cellular mechanisms activated by EPO and IGF-I that protect neurons from apoptosis or axonal/dendritic loss after hypoxiWischemia. HypoxicAschemic injury during stroke causes oxidative and nitrosative stress and produces neuronal damage mediated by increases in glutamate receptor activity and subsequent generation of the free radicals, including nitric oxide (NO) and superoxide (02-). NO and superoxide react to form the neurotoxic product peroxynitrite (ONOO). We show that PI-3 kinase-Akt-1 interaction might be a crucial element in the synergistic effect of EPO and IGF-I. Identification of the neuroprotective signaling pathway(s) for EPO and IGF-I, as proposed in this project, will provide novel strategies to ameliorate loss of neuronal function after stroke and provide an impetus to design new combinatorial therapeutic interventions. Our specific aims are: 1. To characterize EPO- and IGF-I-induced activation of PI-3 kinase that contributes to neuroprotection. 2. To investigate the potential role of Akt in EPO and IGF-I activated neuroprotective signaling in vitro. 3. To study the acute treatment of stroke induced by transient middle artery occlusion (TMCAO) in the mouse : with combined application of EPO and IGF-I. EPO is approved to increase red blood cell mass by the U.S. Food and Drug Administration (FDA) for use in patients having chronic renal failure, receiving chemotherapy for cancer, or with antiretroviral drugs for AIDS. IGF-I is currently in advanced clinical trials for other indications. FDA approval for these other indications could expedite the use of EPOAGF-I for the acute treatment of stroke and other neurodegenerative diseases.